Androgen receptor gene polymorphism analysis: application to screening of heterozygote and to prenatal diagnosis of androgen insensitivity syndrome]

Androgen insensitivity syndromes are X-linked disorders. Restriction fragment length polymorphism analysis of the androgen receptor gene showed that deletions were infrequent. Some mutations have been described. In these conditions, in high-risk family, carrier diagnosis is impossible unless identification of mutations is made. It is thus necessary to detect androgen receptor gene polymorphism in order to differentiate the two maternal X chromosomes. Two androgen receptor gene polymorphisms have been reported (Hind III and exon 1). In this study we analyzed these two gene polymorphisms to detect carriers in at-risk families. The combined results of the two analyses allowed us to detect carriers in 45% of the studied families. In two families the prenatal diagnosis of androgen insensitivity syndrome was performed.     

A statistical approach for detecting genomic aberrations in heterogeneous tumor samples from single nucleotide polymorphism genotyping data

We describe a statistical method for the characterization of genomic aberrations in single nucleotide polymorphism microarray data acquired from cancer genomes. Our approach allows us to model the joint effect of polyploidy, normal DNA contamination and intra-tumour heterogeneity within a single unified Bayesian framework. We demonstrate the efficacy of our method on numerous datasets including laboratory generated mixtures of normal-cancer cell lines and real primary tumours.     

Increased peripheral aromatase activity in prepubertal children with partial androgen insensitivity syndrome

It has been suggested that rate of estrogen formation was higher in patients with androgen insensitivity syndrome (AIS). This work was designed to find out if peripheral aromatase activity could be related to a defect in androgen action in prepubertal children with male pseudohermaphroditism. Fibroblast estrogen production was assayed by a highly specific enzymatic determination. Foreskin fibroblast strains were raised from 17 children with partial androgen insensitivity (PAIS) as defined by dihydrotestosterone binding activity in cells. Results are expressed as pmol estrogens/mg proteins synthetized/day when cultured fibroblasts are incubated with D4-androstenedione. In normal prepubertal boys (n = 19), aromatase activity ranged between 5 and 10 pmol estrogens/mg proteins/day, while in postpubertal boys it varied between 15 and 34 pmol estrogens/mg proteins/day. In prepubertal boys with PAIS (n = 17) aromatase activity is highly elevated: 19.4 +/- 8.4 pmol/mg proteins/day. These results show that (a) peripheral aromatase activity is low before puberty and (b) fibroblast estrogen synthesis is significantly (p less than 0.001) increased in prepubertal children with PAIS. Our data suggest that low utilization of androgens by target cells stimulates the production of estrogen. Peripheral aromatase activity can thus be considered as a 'marker' of androgen insensitivity in prepubertal children with male pseudohermaphroditism.     

Optimization of methods for peripheral blood mononuclear cells isolation and expansion of human gamma delta T cells

Human Vg9/Vδ2 T cells (γδ T cells) are immune surveillance cells both in innate and adaptive immunity and are a possible target for anticancer therapies, which can induce immune responses in a variety of cancers. Small non-peptide antigens such as zoledronate can do activation and expansion of T cells in vitro. It is evident that for adoptive cancer therapies, large numbers of functional cells are needed into cancer patients. Hence, optimization of methods needs to be carried out for the efficient expansion of these T cells. Standardization of peripheral blood mononuclear cells (PBMCs) isolation was devised. Cytokines (interleukin 2 (IL-2) and interleukin 15 (IL-15)) and zoledronate were also standardized for different concentrations. It was found that an increased number of PBMCs were recovered when washing was done at 1100 revolution per minute (rpm). Significantly high expansion fold was (2524 ± 787 expansion fold) achieved when stimulation of PBMCs was done with 1 µM of zoledronate and both cytokines IL-2 and IL-15 supported the expansion and survival of cells at the concentrations of 100 IU/ml and 10 ng/ml respectively. 14-day cultures showed highly pure (91.6 ± 5.1%) and live (96.5 ± 2.5%) expanded γδ T cells. This study aimed to standardize an easy to manipulate technique for the expansion of γδ T cells, giving a higher yield.     

Management of hyperandrogenism in adolescent girls.

Hyperandrogenism in adolescent girls can be a troubling problem because of the difficulty in establishing a diagnosis and in prescribing appropriate therapy. Androgen excess in adolescent patients encompasses a spectrum of clinical presentations, including acne, hirsutism, oligomenorrhea, amenorrhea, virilism, and ovarian cysts. Androgen excess is a clinical and chemical feature of idiopathic hirsutism, late-onset forms of congenital adrenal hyperplasia, and polycystic ovarian disease; in some cases, functional hyperandrogenism is discussed. We recommend screening for hyperandrogenism by measuring blood levels of testosterone, dehydroepiandrosterone sulfate, and delta 4-androstenedione, while others propose a first dexamethasone suppression test for evaluation of free testosterone, dehydroepiandrosterone sulfate, and cortisol. Treatment will be chosen according to particular symptoms such as acne, hirsutism, obesity, or oligomenorrhea.     

How Subchronic and Chronic Health Effects can be Neglected for GMOs, Pesticides or Chemicals

Chronic health effects are increasing in the world such as cancers, hormonal, reproductive, nervous, or immune diseases, even in young people. During regulatory toxicological subchronic tests to prevent these on mammalian health, prior commercialization of chemicals, including pesticides and drugs, or GMOs, some statistically significant findings may be revealed. This discussion is about the need to investigate the relevant criteria to consider those as biologically significant. The sex differences and the non linear dose or time related effects should be considered in contrast to the claims of a Monsanto-supported expert panel about a GMO, the MON 863 Bt maize, but also for pesticides or drugs, in particular to reveal hormone-dependent diseases and first signs of toxicities.     

Malaria Parasite Invasion of the Mosquito Salivary Gland Requires Interaction between the Plasmodium TRAP and the Anopheles Saglin Proteins

SM1 is a twelve-amino-acid peptide that binds tightly to the Anopheles salivary gland and inhibits its invasion by Plasmodium sporozoites. By use of UV-crosslinking experiments between the peptide and its salivary gland target protein, we have identified the Anopheles salivary protein, saglin, as the receptor for SM1. Furthermore, by use of an anti-SM1 antibody, we have determined that the peptide is a mimotope of the Plasmodium sporozoite Thrombospondin Related Anonymous Protein (TRAP). TRAP binds to saglin with high specificity. Point mutations in TRAP''s binding domain A abrogate binding, and binding is competed for by the SM1 peptide. Importantly, in vivo down-regulation of saglin expression results in strong inhibition of salivary gland invasion. Together, the results suggest that saglin/TRAP interaction is crucial for salivary gland invasion by Plasmodium sporozoites.     

Design,synthesis and anti-melanogenic effect of cinnamamide derivatives

Pigmentation disorders are attributed to excessive melanin which can be produced by tyrosinase. Therefore, tyrosinase is supposed to be a vital target for the treatment of disorders associated with overpigmentation. Based on our previous findings that an (E)-β-phenyl-α,β-unsaturated carbonyl scaffold can play a key role in the inhibition of tyrosinase activity, and the fact that cinnamic acid is a safe natural substance with a scaffolded structure, it was speculated that appropriate cinnamic acid derivatives may exhibit potent tyrosinase inhibitory activity. Thus, ten cinnamamides were designed, and synthesized by using a Horner-Emmons olefination as the key step. Cinnamamides 4 (93.72% inhibition), 9 (78.97% inhibition), and 10 (59.09% inhibition) with either a 2,4-dihydroxyphenyl, or 4-hydroxy-3-methoxyphenyl substituent showed much higher mushroom tyrosinase inhibition at 25?µM than kojic acid (18.81% inhibition), used as a positive control. Especially, the two cinnamamides 4 and 9 having a 2,4-dihydroxyphenyl group showed the strongest inhibition. Docking simulation with tyrosinase revealed that these three cinnamamides, 4, 9, and 10, bind to the active site of tyrosinase more strongly than kojic acid. Cell-based experiments carried out using B16F10 murine skin melanoma cells demonstrated that all three cinnamamides effectively inhibited cellular tyrosinase activity and melanin production in the cells without cytotoxicity. There was a close correlation between cellular tyrosinase activity and melanin content, indicating that the inhibitory effect of the three cinnamamides on melanin production is mainly attributed to their capability for cellular tyrosinase inhibition. These results imply that cinnamamides having the (E)-β-phenyl-α,β-unsaturated carbonyl scaffolds are promising candidates for skin-lighting agents.     

Niosomes for oral delivery of nateglinide: in situ–in vivo correlation

Niosomes have been claimed to enhance intestinal absorption and to widen the absorption window of acidic drugs. This was reported after monitoring the intestinal absorption in situ. Accordingly, the aim of this work was to investigate the effect of niosomal encapsulation on intestinal absorption and oral bioavailability of nateglinide. This was conducted with the goal of correlation between in situ intestinal absorption and in vivo availability. The drug was encapsulated into proniosomes. The niosomes resulting after hydration of proniosomes were characterized with respect to vesicle size and drug entrapment efficiency. The in situ rabbit intestinal absorption of nateglinide was monitored from its aqueous solution and niosomes. Streptozotocin was used to induce diabetes in albino rats which were then used to assess the hypoglycemic effect of nateglinide after oral administration of aqueous dispersion and niosomal systems. The prepared vesicles were in the nanoscale with the recorded size being 283?nm. The entrapment efficiency depended on the pH of the formulation. The in situ intestinal absorption reflected non-significant alteration in the membrane transport parameters of the drug after niosomal encapsulation compared with the free drug solution. In contrast, niosomes showed significant improvement in the rate and extent of the hypoglycemic effect compared with the unprocessed drug. This discrepancy can be attributed to different transport pathway for the drug after niosomal inclusion with the vesicles undergoing translymphatic transport which can minimize presystemic metabolism. However, this requires confirmatory investigations. In conclusion niosomes can enhance oral bioavailability of nateglinide with the absorption being through nontraditional pathway.